Metagenomic Profiling of Internationally Sourced Sewage Influents and Effluents Yields Insight into Selecting Targets for Antibiotic Resistance Monitoring

It has been debated whether wastewater treatment plants
(WWTPs)
primarily act to attenuate or amplify antibiotic resistance genes
(ARGs). However, ARGs are highly diverse with respect to their resistance
mechanisms, mobilities, and taxonomic hosts and therefore their behavior
in WWTPs should not be expected to be universally conserved. We applied
metagenomic sequencing to wastewater influent and effluent samples
from 12 international WWTPs to classify the behavior of specific ARGs
entering and exiting WWTPs. In total, 1079 different ARGs originating
from a variety of bacteria were detected. This included ARGs that
could be mapped to assembled scaffolds corresponding to nine human
pathogens. While the relative abundance (per 16S rRNA gene) of ARGs
decreased during treatment at 11 of the 12 WWTPs sampled and absolute
abundance (per mL) decreased at all 12 WWTPs, increases in relative
abundance were observed for 40% of the ARGs detected at the 12th WWTP.
Also, the relative abundance of mobile genetic elements (MGE) increased
during treatment, but the fraction of ARGs known to be transmissible
between species decreased, thus demonstrating that increased MGE prevalence
may not be generally indicative of an increase in ARGs. A distinct
conserved resistome was documented in both influent and effluent across
samples, suggesting that well-functioning WWTPs generally attenuate
influent antibiotic resistance loads. This work helps inform strategies
for wastewater surveillance of antibiotic resistance, highlighting
the utility of tracking ARGs as indicators of treatment performance
and relative risk reduction.